Altered polypeptides, immunoconjugates thereof, and methods related thereto

Abstract

The present invention features inter alia altered binding polypeptides having engineered cysteine residues or analogs thereof at a predetermined site within, for example, a constant region domain or a portion thereof. The engineered cysteine residues or analogs thereof provide sites for conjugating effector moieties (e.g. diagnostic or therapeutic agents) that impart novel functionality to the binding polypeptide, preferably without interfering with a desirable property (e.g. an Fc-mediated effector function). The invention includes methods for the rational design of such altered polypeptides, as well as methods for modifying (ie. conjugating) the altered polypeptides with desirable effector moieties. Particular modified binding polypeptides (ie. immunoconjugates) of altered binding polypeptides and methods for utilizing such modified binding polypeptides as protein-based therapeutics are also provided.

Description

[0001]This application is a continuation application of International Patent Application No. PCT / US2006 / 30234, filed Aug. 1, 2006, which claims the benefit of priority to U.S. provisional patent application No. 60 / 704,603, filed on Aug. 1, 2005, the content of which is hereby incorporated by reference in its entirety.[0002]This application is also related to U.S. Provisional Application Ser. Nos. 60 / 592,886 and 60 / 592,787, both entitled “Binding Molecules and Methods of Use Thereof,” and both filed on Jul. 30, 2005. This application is also related to PCT / US05 / 27262, titled “Binding Molecules and Methods of Use Thereof,” filed on Aug. 1, 2005. The contents of these applications are incorporated in their entirety by this reference.BACKGROUND OF THE INVENTION[0003]Conjugation is a useful method for engineering novel functional properties in polypeptides. Conventional conjugation methods for modifying polypeptides with functional moieties, however, have certain shortcomings. First, the...

AI Technical Summary

Benefits of technology

[0047]In one aspect, the invention pertains to an altered polypeptide comprising at least one CH3 domain or portion thereof wherein the CH3 domain comprises an engineered cysteine or thiol-containing analog thereof at one or more amino acid positions selected from the group consisting of amino acid positions 350, 352, 353, 355, 358-366, 368, 370, 371, 389-391, 394-396, 398,400-407, 409-423, 4

Problems solved by technology

First, the chemistry of the cross-linking reaction is usually very inefficient.

As a result, a significant amount of unmodified polypeptide is lost during the manufacturing.

Second, the modified polypeptide produced by chemical cross-linking contains a chemical cross-linker fragment that can be immunogenic or toxic.

This is especially important in therapeutic applications where the modified polypeptide may have to be administered many times. Third, many cross-linking methods are neither site-specific nor stereo-specific, and consequently, may decrease the functionality, specificity, affinity, or avidity of the polypeptide.

Fourth, the cross-linking method may not allow for precise stoichiometric control of the number of modifications to the polypeptide.

A heterogeneous mixture of polypeptides so modified is inadequate for diagnostic and therapeutic use as it is inadequately defined and consists of a population of different chemical entities with varying biological properties.

However, these methods are also problematic, as disulfide bonds formed by native cysteines are usually essential for maintaining the structure and function of the polypeptide.

Unfortunately, however, the engineered cysteine residue will often result in drastic effects on the favorable activities (e.g. effector function) that constant region sequences provide.

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